The identification of cellular DNA sequences determining tumorigenicity has relied almost solely on the transfection assay for production of foci in NIH3T3 cells. This method raises serious questions regarding the importance of such isolated genes in human systems and it may also be greatly restricted in the type of transforming genes it can detect. We are developing a system utilizing a more appropriate human cell line as recipient in transfection and to use the transfected cells in both focus and tumor formation assays. The first objective was the identification of a human cell line which was an efficient recipient for DNA transfer and which was non-tumorigenic in the athymic nude mouse without being immunologically rejected. Certain hybrids of human fibroblasts and the normally tumorigenic HeLa cell line fulfill these criteria, particularly as they retain a cryptic tumorigenic potential and are theoretically capable of being rendered tumorigenic by one or a small number of further changes. To obtain a positive control, these lines are being tested for focus and tumor formation following transfection with cloned viral onc genes. The parents of the hybrids, the hybrids and their rare tumorigenic derivatives are being examined for mRNA changes using viral onc gene probes to identify the mechanism of suppression of tumorigenicity in the hybrids.